1. Field of the Invention
A method to decrease the gastro-intestinal bacterial load of animals by feeding a diet or solution containing terpenes and/or liposome-terpene combination 1 to 25 days before slaughter.
2. Discussion of the Background
In a normal, steady-state eating pattern, the intestine of animals is usually empty 4 to 6 hours after a feeding. For this reason, feed is withdrawn 4 to 12 hours before slaughter. This procedure is not without compromises. Because of the lack of feed, the animal loses weight and the intestinal wall becomes fragile and prone to rupture. This results in spilling of the intestinal content during slaughter and evisceration (Buhr et al, 1998). In the case of poultry, spilling causes contamination with whatever bacteria is present. Another problem with feed withdrawal is the increased number of animals that will practice coprophagy (eating of feces), resulting in higher bacterial counts in the intestinal contents and the final processed products (meat) (Byrd et al, 1998). When the feed is not withdrawn, the crop and intestinal tract are full of feed, which also results in breaking and spilling of the contents. Contamination can also originate from bacteria present in feathers and skin. Feed withdrawal may reduce the contamination of carcasses but it also results in a decline of volatile fatty acids and a resultant increase in cecal pH which is correlated with an increase in fecal shedding of foodborne pathogens.
There are not any safe procedures or products that can decrease carcass contamination or decrease the loss in animal weight. There are chemicals that recently have shown promising results. The positive results from the use of terpenes have been reported.
Terpenes (pinene, nerol, citral, menthol, d-limonene and others) are widespread in nature, mainly in plants as constituents of essential oils. Their building block is the hydrocarbon isoprene (C5H8)n. Terpenes are effective, nontoxic dietary antitumor agents which act through a variety of mechanisms of action and hold promise as a novel class of antitumor drugs for human cancer (Crowell and Gould, 1994 and Crowell et al, 1996). Geraniol, tocotrienol, perillyl alcohol, b-ionone and d-limonene suppress hepatic HMG-COA reductase activity, a rate limiting step in cholesterol synthesis, and modestly lower cholesterol levels in animals (Elson and Yu, 1994). D-limonene and geraniol reduced mammary tumors (Elegbede et al , 1984 and 1986 and Karlson et al, 1996) or suppressed the growth of transplanted tumors(Yu et al, 1995).
Terpenes (citral, geraniol, eugenol, menthol, cinnamic aldehyde) have also been found to inhibit the in-vitro growth of bacteria and fungi (Chaumont and Leger, 1992, Moleyar and Narasimham, 1992 and Pattnaik, et al 1997) and some internal and external parasites (Hooser, et al, 1986). Geraniol was found to inhibit growth of candida albicans and saccharomyces cerevisiae strains by enhancing the rate of potassium leakage disrupting membrane fluidity (Bard, et al, 1988). B-ionone has antifungal activity which was determined by inhibition of spore germination, and growth inhibition in agar (Mikhlin et al, 1983 and Salt et al, 1986). Teprenone has an antibacterial effect on H. pylori (Ishii, 1993). Solutions of 11 different terpenes were effective in inhibiting the growth of pathogenic bacteria in in-vitro tests; levels ranging between 100 ppm and 1000 ppm were effective. The terpenes were diluted in water with 1% tween 20 (Kim et al, 1995).
There may be different modes of action of terpenes against bacteria. They could (1)interfere with the phospholipid bilayer of the cell membrane (2)impair a variety of enzyme systems (HMG-reductase) and (3) destroy or inactivate genetic material.
The addition of terpenes to the diet or to the drinking water would reduce the bacterial load, not only in feed and water, but also in the intestine. Pathogenic bacteria like salmonella, E. coli and listeria would be decreased, thereby reducing the chance of contamination and the risk of pathogenic microorganisms in the final edible product.
A method for the reduction of the gastrointestinal bacterial load by feeding a diet or solution containing a terpene, a terpene mixture or a liposome-terpene combination.
Terpenes, which are GRAS (Generally Recognized As Safe) have been found to inhibit the growth of cancerous cells, decrease tumor size, decrease cholesterol level and have a biocidal effect on microorganisms in vitro. Onawunmi (1989) showed that media with more than 0.01% citral reduced the concentration of E. coli and at 0.08% there was a bactericidal effect. Barranx, et al(1998) teach us a terpene formulation, based on pine oil, used as disinfectant or antiseptic cleaner. Koga, et al (1998) teach that a terpene found in rice has antifungal activity. Neither of them suggested the use of a terpene or terpene mixture for the internal disinfection of the gastrointestinal tract of animals by the addition of a terpene, a terpene mixture or a liposome-terpene combination into feed or water.
Several U.S. Patents (U.S. Pat. No. 5,547,677, U.S. Pat. No. 5,549,901, U.S. Pat. No. 5,618,840, U.S. Pat. No. 5,629,021, U.S. Pat. No. 5,662,957, U.S. Pat. No. 5,700,679, U.S. Pat. No. 5,730,989)teach us that certain types of oil-in-water emulsions have antimicrobial, adjuvant and delivery properties. By encapsulating terpenes within these emulsions or mixing them together, the antimicrobial effect will be increased, i.e. (1) the liposome will disrupt the bacterial membrane and (2) the terpenes will be more effective in disrupting cytoplasmatic enzymes.
Some procedures are used in order to change the microflora of animals, but they will not alter bacteria number. Competitive Exclusion (CE) has been utilized to decrease the incidence of pathogenic organism in animals. CE will create a competition for nutrients between good bacteria and pathogenic bacteria, but it is unlikely that the usage of beneficial bacteria can significantly reduce the bacteria count in the final processed product. The animal industry feeds low levels of antibiotics to decrease bacterial exposure, which results in an improvement in health and performance, but does not eliminate great numbers of bacteria. Currently federal agencies have approved the use of low level radiation to kill bacteria in meat, but the use of radiation at any level will always be challenged by concerned groups.
The short use or continuous use of a terpene mixture or liposome-terpene composition will have the same effect as antibiotic feeding, with the advantage that terpenes will not result in resistant bacteria or drug residues in animal produce. Short term or continuous terpene feeding can be obtained by adding the terpene, terpene mixture or liposome-terpene composition in the feed or in the drinking water of animals. The terpenes can also be delivered encapsulated within liposomes.